Microbicidal composition

ABSTRACT

A synergistic microbicidal composition having two components. The first component is a nonionic surfactant with structure: R 2 O(CH 2 CH(CH 3 )O) 3 (CH 2 CH 2 O) 5 H, where R 2  is a mixture of C 8 -C 14  linear alkyl groups. The second component is 2-bromo-2-nitropropane-1,3-diol. The weight ratio of the nonionic surfactant to 2-bromo-2-nitropropane-1,3-diol is from 1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286.

This invention relates to microbicidal compositions containing2-bromo-2-nitropropane-1,3-diol (BNPD),tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione (DAZOMET) orBIOBAN P-1487 (ca. 81:5 mixture of 4-(2-nitrobutyl)morpholine and4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine, respectively) and asurfactant.

A composition containing 5-chloro-2-methylisothiazolin-3-one,2-methylisothiazolin-3-one and a nonionic dispersant is disclosed inU.S. Pat. No. 4,295,932. The composition contains a 3:1 mixture of5-chloro-2-methylisothiazolin-3-one and 2-methylisothiazolin-3-one, anda copolymer of ethylene oxide and propylene oxide which appears to havethe same composition as PLURONIC L61 or TERGITOL L61 dispersant.However, there is a need for combinations of microbicides havingsynergistic activity against various strains of microorganisms toprovide effective control of the microorganisms. Moreover, there is aneed for such combinations containing lower levels of individualmicrobicides for environmental and economic benefit. The problemaddressed by this invention is to provide such synergistic combinationsof microbicides.

STATEMENT OF THE INVENTION

The present invention is directed to a synergistic microbicidalcomposition comprising: (a) a nonionic surfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅Hwhere R² is a mixture of C₈-C₁₄ linear alkyl groups; and (b)2-bromo-2-nitropropane-1,3-diol; wherein a weight ratio of said nonionicsurfactant to 2-bromo-2-nitropropane-1,3-diol is from 1:0.08 to 1:0.12or 1:0.2 to 1:1.8286.

The present invention is further directed to a synergistic microbicidalcomposition comprising: (a) a nonionic surfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅Hwhere R² is a mixture of C₈-C₁₄ linear alkyl groups; and (b)tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione; wherein a weightratio of said nonionic surfactant totetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione is from 1:0.0024to 1:0.4571.

The present invention is further directed to a synergistic microbicidalcomposition comprising: (a) a nonionic surfactant with structure:R¹O(CH₂CH(CH₃)O)₅(CH₂CH₂O)₉Hwhere R¹ is a C₈ alkyl group; and (b) a mixture of4-(2-nitrobutyl)morpholine and4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine in a weight ratio of81:5; wherein a weight ratio of said nonionic surfactant to said mixtureis from 1:0.0006 to 1:0.2743.

The present invention is further directed to methods for inhibiting thegrowth of microorganisms in aqueous media by adding to an aqueous mediuma nonionic surfactant as described herein and2-bromo-2-nitropropane-1,3-diol,tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione or a ca. 81:5mixture of 4-(2-nitrobutyl)morpholine and4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine in the ratios describedherein.

DETAILED DESCRIPTION OF THE INVENTION

“BNPD” is 2-bromo-2-nitropropane-1,3-diol, “DAZOMET” has as an activeingredient tetrahydro-3,5-dimethyl-2H-1,3,5-thiadiazine-2-thione (CASNo. 533-74-4) and “BIOBAN P-1487” is an approximately 81:5 (weight)mixture of 4-(2-nitrobutyl)morpholine and4,4′-(2-ethyl-2-nitrotrimethylene)dimorpholine. As used herein, thefollowing terms have the designated definitions, unless the contextclearly indicates otherwise. The term “microbicide” refers to a compoundcapable of inhibiting the growth of or controlling the growth ofmicroorganisms; microbicides include bactericides, fungicides andalgaecides. The term “microorganism” includes, for example, fungi (suchas yeast and mold), bacteria and algae. The following abbreviations areused throughout the specification: ppm=parts per million by weight(weight/weight), mL=milliliter. Unless otherwise specified, temperaturesare in degrees centigrade (° C.), references to percentages arepercentages by weight (wt %) and amounts and ratios are on an activeingredient basis, i.e., total weight of BNPD, DAZOMET or BIOBAN P-1487active ingredients and the nonionic surfactant. Numbers of polymerizedunits of propylene oxide or ethylene oxide are number averages.

Preferably, the weight ratio of the nonionic surfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅Hwhere R² is a mixture of C₈-C₁₄ linear alkyl groups to BNPD is from1:0.2286 to 1:1.18286.

The present invention is further directed to a method for inhibiting thegrowth of mold, preferably C. albicans, in an aqueous medium by adding:(a) a nonionic surfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅Hwhere R² is a mixture of C₈-C₁₄ linear alkyl groups; and (b) BNPD;wherein a weight ratio of said nonionic surfactant to BNPD is from1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286, preferably from 1:0.2286 to1:1.18286.

Preferably, the weight ratio of the nonionic surfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅Hwhere R² is a mixture of C₈-C₁₄ linear alkyl groups to DAZOMET is from1:0.0114 to 1:0.4571.

The present invention is further directed to a method for inhibiting thegrowth of S. aureus, in an aqueous medium by adding: (a) a nonionicsurfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅Hwhere R² is a mixture of C₈-C₁₄ linear alkyl groups; and (b) DAZOMET;wherein a weight ratio of said nonionic surfactant to DAZOMET is from1:0.0114 to 1:0.4571.

Preferably, the weight ratio of the nonionic surfactant with structure:R¹O(CH₂CH(CH₃)O)₅(CH₂CH₂O)₉Hwhere R¹ is a C₈ alkyl group to BIOBAN P-1487 is from 1:0.0160 to1:0.2743.

The present invention is further directed to a method for inhibiting thegrowth of fungi, preferably A. niger, in an aqueous medium by adding:(a) a nonionic surfactant with structure:R¹O(CH₂CH(CH₃)O)₅(CH₂CH₂O)₉Hwhere R¹ is a C₈ alkyl group to BIOBAN P-1487 is from 1:0.0160 to1:0.2743.

R² is a mixture of C₈-C₁₄ linear alkyl groups. Preferably, the C₈-C₁₄linear alkyl groups comprise from 50 to 85 wt % C₈-C₁₀ linear alkylgroups and 15 to 50 wt % C₁₂-C₁₄ linear alkyl groups, preferably from 60to 75 wt % C₈-C₁₀ linear alkyl groups and 25 to 40 wt % C₁₂-C₁₄ linearalkyl groups, preferably about 70 wt % C₈-C₁₀ linear alkyl groups andabout 30 wt % C₁₂-C₁₄ linear alkyl groups. Preferably, the linear alkylgroups are derived from seed oil. Preferably, R¹ is 2-ethylhexyl.

Preferably, each of the compositions is substantially free ofmicrobicides other than the nonionic surfactant and BNPD, DAZOMET orBIOBAN P-1487, i.e., it has less than 1 wt % of microbicides other thanthe nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487 based ontotal weight of active ingredients, preferably less than 0.5 wt %,preferably less than 0.2 wt %, preferably less than 0.1 wt %.Preferably, when the nonionic surfactant and BNPD, DAZOMET or BIOBANP-1487 are added to an aqueous medium, the medium is substantially freeof other microbicides, i.e., it has less than 1 wt % of microbicidesother than the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487based on total weight of active ingredients, preferably less than 0.5 wt%, preferably less than 0.2 wt %, preferably less than 0.1 wt %.

The compositions of this invention may contain other ingredients, e.g.,defoamers and emulsifiers. The microbicidal compositions of the presentinvention can be used to inhibit the growth of microorganisms or higherforms of aquatic life (such as protozoans, invertebrates, bryozoans,dinoflagellates, crustaceans, mollusks, etc) by introducing amicrobicidally effective amount of the compositions into an aqueousmedium subject to microbial attack. Suitable aqueous media are found in,for example: industrial process water; electrocoat deposition systems;cooling towers; air washers; gas scrubbers; mineral slurries; wastewatertreatment; ornamental fountains; reverse osmosis filtration;ultrafiltration; ballast water; evaporative condensers; heat exchangers;pulp and paper processing fluids and additives; starch; plastics;emulsions; dispersions; paints; latices; coatings, such as varnishes;construction products, such as mastics, caulks, and sealants;construction adhesives, such as ceramic adhesives, carpet backingadhesives, and laminating adhesives; industrial or consumer adhesives;photographic chemicals; printing fluids; household products, such asbathroom and kitchen cleaners; cosmetics; toiletries; shampoos; soaps;personal care products such as wipes, lotions, sunscreen, conditioners,creams, and other leave-on applications; detergents; industrialcleaners; floor polishes; laundry rinse water; metalworking fluids;conveyor lubricants; hydraulic fluids; leather and leather products;textiles; textile products; wood and wood products, such as plywood,chipboard, flakeboard, laminated beams, oriented strandboard, hardboard,and particleboard; petroleum processing fluids; fuel; oilfield fluids,such as injection water, fracture fluids, and drilling muds; agricultureadjuvant preservation; surfactant preservation; medical devices;diagnostic reagent preservation; food preservation, such as plastic orpaper food wrap; food, beverage, and industrial process pasteurizers;toilet bowls; recreational water; pools; and spas.

The specific amount of the microbicidal compositions of this inventionnecessary to inhibit or control the growth of microorganisms in anapplication will vary. Amounts of the compositions of the presentinvention are given on the basis of active ingredients (i.e., nonionicsurfactant and BNPD, DAZOMET or BIOBAN P-1487). For BNPD/surfactant usedin paper applications, preferably the concentration is 2 to 50 ppm,preferably 5 to 40 ppm. For BNPD/surfactant used in oil and gas drillingapplications, preferably the concentration is 10 to 500 ppm, preferably20 to 350 ppm. For BNPD/surfactant used in metalworking fluids,preferably the concentration is 150 to 2000 ppm, preferably 300 to 1500ppm. For BIOBAN P-1487/surfactant used in metalworking fluids,preferably the concentration is 600 to 1800 ppm, preferably 800 to 1500ppm. For BIOBAN P-1487/surfactant used in fuel applications, preferablythe concentration is 300 to 4000 ppm, preferably 500 to 3000 ppm. Fuelapplications include the preservation of hydrocarbons for fuel, e.g.,bioethanol applications, diesel storage tank preservation and generalfuel tank preservation. In a method of this invention, a composition istreated to inhibit microbial growth by adding, together or separately,the nonionic surfactant and BNPD, DAZOMET or BIOBAN P-1487, in amountsthat would produce the concentrations indicated above.

EXAMPLES

Surfactants and biocides were evaluated for synergy by determining thesynergy index (S.I.) of the combination. Synergy index was calculatedbased on minimum inhibitory concentrations (MIC) of two antimicrobialcompounds (A and B) alone and in combinations. The tests organisms wereGram negative bacteria (Pseudomonas aeruginosa ATCC #15442), Grampositive bacteria (Staphylococcus aureus ATCC #6538), yeast (Candidaalbicans ATCC #10203) and mold (Aspergillus niger ATCC #16404). Contacttime for the bacteria was 24 and 48 hours, yeast was 48 and 72 hrs, and3 and 7 days for mold. The test was carried out in 96 well microtiterplates.

-   Surf. A R¹O(CH₂CH(CH₃)O)₅(CH₂CH₂O)₉H, where R¹ is 2-ethylhexyl-   Surf. D R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅H-   Surf. E R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₇H    In Surf. D and Surf. E, R² is a mixture of C₈-C₁₄ linear alkyl    groups (70% C₈-C₁₀ linear alkyl and 30% C₁₂-C₁₄ linear alkyl)    Inoculums Used

Inoculum Size of organisms (CFU/ml) Staphylococcus PseudomonasAspergillus Candida aureus aeruginosa niger albicans ATCC# ATCC# ATCC#ATCC# Surfactants 6538 15442 16404 10203 Surf. A 1.156E+06 8.134E+071.156E+06  1.156+06 Surf. D 1.808E+05 1.156E+08 1.156E+06 5.726E+05Surf. E 1.808E+06 5.727E+07 5.726E+05 1.808E+06

TABLE 4 Media Used Media Used for testing Staphylococcus PseudomonasAspergillus Candida aureus aeruginosa niger albicans ATCC# ATCC# ATCC#ATCC# 6538 15442 16404 10203 10% Tryptic soy 10% Tryptic soy Potatodextrose Potato dextrose broth broth broth broth

The pH of the Triptic soy broth was 7.3 and the Potato dextrose brothwas 5.1.

The test results for demonstration of synergy of the MIC combinationsare shown in the tables below. Each table shows the results forcombinations of two components against the microorganisms tested withincubation times; the end-point activity in ppm measured by the MIC forcompound A alone (CA), for component B alone (CB), and the mixture (Ca)and (Cb); the calculated SI value; and the range of synergistic ratiosfor each combination tested. SI is calculated as follows:Ca/CA+Cb/CB=Synergy Index(“SI”)

Wherein:

-   -   CA=concentration of compound A in ppm, acting alone, which        produced an end point (MIC of Compound A).    -   Ca=concentration of compound A in ppm, in the mixture, which        produced an end point.    -   CB=concentration of compound B in ppm, acting alone, which        produced an end point (MIC of Compound B).    -   Cb=concentration of compound B in ppm, in the mixture, which        produced an end point.        When the sum of Ca/CA and Cb/CB is greater than one, antagonism        is indicated. When the sum is equal to one, additivity is        indicated, and when less than one, synergism is demonstrated.

The ratio ranges at which BNPD, DAZOMET or BIOBAN P-1487 and thesurfactants were tested are as summarized in the following tables:

BNPD with Surf. A Organism ATCC # From To Ratio Range Staphylococcus6538 0.02:20,000  100:218.75 1:0.000001- aureus 1:0.45714 Aspergillus16404  0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714 Candida 10203 0.2:20,000 1,000:218.75 1:0.00001- albicans 1:4.5714 Pseudomonas 154420.02:20,000  100:218.75 1:0.000001- aeruginosa 1:0.45714

BNPD with Surf. E Organism ATCC# From To Ratio Range Staphylococcus 65380.02:20,000  100:218.75 1:0.000001- aureus 1:0.45714 Aspergillus 16404 0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714 Candida 10203 0.2:20,000 1,000:218.75 1:0.00001- albicans 1:4.5714 Pseudomonas 154420.02:20,000  100:218.75 1:0.000001- aeruginosa 1:0.45714

BNPD with Surf. D Organism ATCC# From To Ratio Range Staphylococcus 65380.02:20,000  100:218.75 1:0.000001- aureus 1:0.45714 Aspergillus 16404 0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714 Candida 10203 0.2:20,000 1,000:218.75 1:0.00001- albicans 1:4.5714 Pseudomonas 154420.02:20,000  100:218.75 1:0.000001- aeruginosa 1:0.45714

BIOBAN P-1487 with Surf. A Organism ATCC# From To Ratio RangeStaphylococcus 6538 0.002:20,000   10:218.75 1:0.0000001- aureus1:0.0457143 Aspergillus 16404 0.02:20,000 100:218.75 1:0.000001- niger1:0.45714 Candida 10203 0.02:20,000 100:218.75 1:0.000001- albicans1:0.45714 Pseudomonas 15442 0.02:20,000 100:218.75 1:0.000001-aeruginosa 1:0.45714

BIOBAN P-1487 with Surf. E Organism ATCC# From To Ratio RangeStaphylococcus 6538 0.02:20,000 100:218.75 1:0.000001- aureus 1:0.45714Aspergillus 16404 0.02:20,000 100:218.75 1:0.000001- niger 1:0.45714Candida 10203 0.02:20,000 100:218.75 1:0.000001- albicans 1:0.45714Pseudomonas 15442 0.02:20,000 100:218.75 1:0.000001- aeruginosa1:0.45714

BIOBAN P-1487 with Surf. D Organism ATCC# From To Ratio RangeStaphylococcus 6538 0.02:20,000  100:218.75 1:0.000001- aureus 1:0.45714Aspergillus 16404  0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714Candida 10203 0.02:20,000  100:218.75 1:0.000001- albicans 1:0.45714Pseudomonas 15442  0.2:20,000 1,000:218.75 1:0.00001- aeruginosa1:4.5714

DAZOMET with Surf. A Organism ATCC# From To Ratio Range Staphylococcus6538 0.2:20,000 1,000:218.75 1:0.00001- aureus 1:4.5714 Aspergillus16404 0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714 Candida 102030.2:20,000 1,000:218.75 1:0.00001- albicans 1:4.5714 Pseudomonas 154420.2:20,000 1,000:218.75 1:0.00001- aeruginosa 1:4.5714

DAZOMET with Surf. E Organism ATCC# From To Ratio Range Staphylococcus6538 0.2:20,000 1,000:218.75 1:0.00001- aureus 1:4.5714 Aspergillus16404 0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714 Candida 102030.2:20,000 1,000:218.75 1:0.00001- albicans 1:4.5714 Pseudomonas 154420.2:20,000 1,000:218.75 1:0.00001- aeruginosa 1:4.5714

DAZOMET with Surf. D Organism ATCC# From To Ratio Range Staphylococcus6538 0.2:20,000 1,000:218.75 1:0.00001- aureus 1:4.5714 Aspergillus16404 0.2:20,000 1,000:218.75 1:0.00001- niger 1:4.5714 Candida 102030.2:20,000 1,000:218.75 1:0.00001- albicans 1:4.5714 Pseudomonas 154420.2:20,000 1,000:218.75 1:0.00001- aeruginosa 1:4.5714

-   A: Surf. A-   B: BNPD-   Media: 1/10 TSB-   Inoculum size: 8.134E+07 CFU/ml

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) Ps. aeruginosa >20000 10 20000 8 <1.80 1:0.0004 ATCC#15442 >20000 10 10000 8 <1.30 1:0.0008 >20000 10 5000 8 <1.051:0.0016 >20000 10 2500 8 <0.93 1:0.0032 >20000 10 1750 8 <0.891:0.0046 >20000 10 875 8 <0.84 1:0.0091 >20000 10 437.5 8 <0.821:0.0183 >20000 10 218.75 8 <0.81 1:0.0366No Synergy: A. niger, C. albicans, S. aureus

-   A: Surf. D-   B: BNPD-   Media: PDB-   Inoculum size: 1.156E+06

PPM AI MIC Values (3rd day) Test Alone Combination Ratio Organism CA CBCa Cb S.I. (Ca:Cb) A. niger >20000 1000 20000 500 <1.50 1:0.0250 ATCC#16404 >20000 1000 10000 400 <0.90 1:0.0400 >20000 1000 10000 500 <1.001:0.0500 >20000 1000 5000 500 <0.75 1:0.1000 >20000 1000 5000 600 <0.851:0.1200 >20000 1000 5000 800 <1.05 1:0.1600 >20000 1000 2500 500 <0.631:0.2000 >20000 1000 2500 600 <0.73 1:0.2400 >20000 1000 2500 800 <0.931:0.3200 >20000 1000 1750 600 <0.69 1:0.3429 >20000 1000 1750 800 <0.891:0.4571 >20000 1000 875 600 <0.64 1:0.6857 >20000 1000 875 800 <0.841:0.9143

-   A: Surf. D-   B: BNPD-   Media: PDB-   Inoculum size: 5.726E+05 CFU/ml

PPM AI MIC Values (48 hrs) Test Alone Combination Ratio Organism CA CBCa Cb S.I. (Ca:Cb) C. albicans >20000 1000 20000 400 <1.40 1:0.0200ATCC# 10203 >20000 1000 10000 400 <0.90 1:0.0400 >20000 1000 10000 500<1.00 1:0.0500 >20000 1000 5000 400 <0.65 1:0.0800 >20000 1000 5000 500<0.75 1:0.1000 >20000 1000 5000 600 <0.85 1:0.1200 >20000 1000 5000 800<1.05 1:0.1600 >20000 1000 2500 400 <0.53 1:0.1600 >20000 1000 2500 600<0.73 1:0.2400 >20000 1000 2500 800 <0.93 1:0.3200 >20000 1000 1750 400<0.49 1:0.2286 >20000 1000 1750 600 <0.69 1:0.3429 >20000 1000 1750 800<0.89 1:0.4571 >20000 1000 875 500 <0.54 1:0.5714 >20000 1000 875 600<0.64 1:0.6857 >20000 1000 875 800 <0.84 1:0.9143 >20000 1000 437.5 800<0.82 1:1.8286

-   A: Surf. D-   B: BNPD-   Media: 1/10 TSB-   Inoculum size: 1.16E+08

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) Ps. aeruginosa >20000 10 875 8 <0.84 1:0.0091 ATCC#15442 >20000 10 437.5 8 <0.82 1:0.0183

-   A: Surf. D-   B: BNPD-   Media: 1/10 TSB-   Inoculum size: 1.808E+05

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) S. aureus >20000 20 20000 4 <1.20 1:0.0002 ATCC#6538 >20000 20 10000 3 <0.65 1:0.0003 >20000 20 10000 6 <0.801:0.0006 >20000 20 10000 8 <0.90 1:0.0008 >20000 20 10000 10 <1.001:0.0010 >20000 20 5000 2 <0.35 1:0.0004 >20000 20 5000 4 <0.451:0.0008 >20000 20 5000 8 <0.65 1:0.0016 >20000 20 5000 10 <0.751:0.0020 >20000 20 5000 2 <0.35 1:0.0004 >20000 20 5000 6 <0.551:0.0012 >20000 20 5000 10 <0.75 1:0.0020 >20000 20 1750 2 <0.191:0.0011 >20000 20 1750 8 <0.49 1:0.0046 >20000 20 1750 10 <0.591:0.0057 >20000 20 875 2 <0.14 1:0.0023 >20000 20 875 4 <0.241:0.0046 >20000 20 875 6 <0.34 1:0.0069 >20000 20 875 10 <0.541:0.0114 >20000 20 437.5 6 <0.32 1:0.0137 >20000 20 437.5 8 <0.421:0.0183 >20000 20 437.5 10 <0.52 1:0.0229 >20000 20 218.75 8 <0.411:0.0366 >20000 20 218.75 10 <0.51 1:0.0457

-   A: Surf. E-   B: BNPD-   Media: PDB-   Inoculum size: 1.808E+06 CFU/ml

PPM AI MIC Values (48 hrs) Test Alone Combination Ratio Organism CA CBCa Cb S.I. (Ca:Cb) C. albicans >20000 1000 20000 200 <1.20 1:0.0100ATCC# 10203 >20000 1000 10000 200 <0.70 1:0.0200 >20000 1000 10000 300<0.80 1:0.0300 >20000 1000 10000 500 <1.00 1:0.0500 >20000 1000 5000 200<0.45 1:0.0400 >20000 1000 5000 400 <0.65 1:0.0800 >20000 1000 5000 600<0.85 1:0.1200 >20000 1000 5000 800 <1.05 1:0.1600 >20000 1000 2500 400<0.53 1:0.1600 >20000 1000 2500 600 <0.73 1:0.2400 >20000 1000 2500 800<0.93 1:0.3200 >20000 1000 1750 500 <0.59 1:0.2857 >20000 1000 1750 800<0.89 1:0.4571 >20000 1000 875 800 <0.84 1:0.9143

-   A: Surf. E-   B: BNPD-   Media: 1/10 TSB-   Inoculum size: 5.727E+07 CFU/ml

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) Ps. aeruginosa >20000 10 5000 8 <1.05 1:0.0016 ATCC#15442 >20000 10 2500 8 <0.93 1:0.0032 >20000 10 1750 8 <0.891:0.0046 >20000 10 875 8 <0.84 1:0.0091 >20000 10 437.5 8 <0.821:0.0183 >20000 10 218.75 8 <0.81 1:0.0366

-   A: Surf. E-   B: BNPD-   Media: 1/10 TSB-   Inoculum size: 1.808E+06 CFU/ml

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) S. aureus >20000 20 20000 8 <1.40 1:0.0004ATCC# >20000 20 10000 8 <0.90 1:0.0008 6538 >20000 20 5000 8 <0.651:0.0016 >20000 20 5000 10 <0.75 1:0.0020 >20000 20 2500 8 <0.531:0.0032 >20000 20 2500 10 <0.63 1:0.0040 >20000 20 1750 8 <0.491:0.0046 >20000 20 1750 10 <0.59 1:0.0057 >20000 20 875 8 <0.441:0.0091 >20000 20 875 10 <0.54 1:0.0114

-   No synergy: A. niger-   A: Surf. A-   B: DAZOMET-   Media: 1/10 TSB-   Inoculum size: 8.13E+07-   S. aureus ATCC 6538 No Synergy-   A: Surf. A-   B: DAZOMET-   Media: PDB-   Inoculum size: 1.156E+06

PPM AI MIC Values (3rd day) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) >20000 60 10000 0.6 <0.51 1:0.0001 A. niger >20000 6010000 0.8 <0.51 1:0.0001 ATCC# 16404 >20000 60 10000 1 <0.521:0.0002 >20000 60 10000 2 <0.53 1:0.0003 >20000 60 10000 3 <0.551:0.0004 >20000 60 10000 4 <0.57 1:0.0005 >20000 60 10000 5 <0.581:0.0006 >20000 60 10000 6 <0.60 1:0.0008 >20000 60 10000 8 <0.631:0.0010 >20000 60 10000 10 <0.67 1:0.0020 >20000 60 10000 20 <0.831:0.0030 >20000 60 10000 30 <1.00 1:0.0040 >20000 60 10000 40 <1.171:0.0050 >20000 60 10000 50 <1.33 1:0.0008 >20000 60 5000 4 <0.321:0.0016 >20000 60 5000 8 <0.38 1:0.0020 >20000 60 5000 10 <0.421:0.0040 >20000 60 5000 20 <0.58 1:0.0060 >20000 60 5000 30 <0.751:0.0080 >20000 60 5000 40 <0.92 1:0.0100 >20000 60 5000 50 <1.081:0.0020 >20000 60 2500 5 <0.21 1:0.0024 >20000 60 2500 6 <0.231:0.0032 >20000 60 2500 8 <0.26 1:0.0040 >20000 60 2500 10 <0.291:0.0080 >20000 60 2500 20 <0.46 1:0.0120 >20000 60 2500 30 <0.631:0.0160 >20000 60 2500 40 <0.79 1:0.0200 >20000 60 2500 50 <0.961:0.0029 >20000 60 1750 5 <0.17 1:0.0020 >20000 60 1750 6 <0.231:0.0024 >20000 60 1750 8 <0.26 1:0.0032 >20000 60 1750 10 <0.291:0.0040 >20000 60 1750 20 <0.46 1:0.0080 >20000 60 1750 30 <0.631:0.0120 >20000 60 1750 40 <0.79 1:0.0160 >20000 60 1750 50 <0.961:0.0200 >20000 60 875 20 <0.17 1:0.0029 >20000 60 875 30 <0.191:0.0034 >20000 60 875 40 <0.22 1:0.0046 >20000 60 875 50 <0.251:0.0057 >20000 60 437.5 30 <0.42 1:0.0114 >20000 60 437.5 40 <0.591:0.0171 >20000 60 437.5 50 <0.75 1:0.0229 >20000 60 218.75 40 <0.921:0.0286 >20000 60 218.75 50 <0.38 1:0.0229

-   A: Surf. A-   B: DAZOMET-   Media: PDB-   Inoculum size: 1.156E+06

PPM AI MIC Values (48 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) C. albicans >20000 50 2500 40 <0.93 1:0.0160 ATCC#10203 >20000 50 1750 40 <0.89 1:0.0229 >20000 50 437.5 40 <0.821:0.0914 >20000 50 218.75 40 <0.81 1:0.1829

-   A: Surf. A-   B: DAZOMET-   Media: 1/10 TSB-   Inoculum size: 8.134E+07

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) Ps. aeruginosa >20000 80 2500 4 <0.18 1:0.0016 ATCC#15442 >20000 80 2500 5 <0.19 1:0.0020 >20000 80 2500 6 <0.201:0.0024 >20000 80 2500 8 <0.23 1:0.0032 >20000 80 2500 10 <0.251:0.0040 >20000 80 2500 20 <0.38 1:0.0080 >20000 80 2500 30 <0.501:0.0120 >20000 80 2500 40 <0.63 1:0.0160 >20000 80 2500 50 <0.751:0.0200 >20000 80 2500 60 <0.88 1:0.0240 >20000 80 1750 6 <0.161:0.0034 >20000 80 1750 8 <0.19 1:0.0046 >20000 80 1750 10 <0.211:0.0057 >20000 80 1750 20 <0.34 1:0.0114 >20000 80 1750 30 <0.461:0.0171 >20000 80 1750 40 <0.59 1:0.0229 >20000 80 1750 50 <0.711:0.0286 >20000 80 1750 60 <0.84 1:0.0343 >20000 80 437.5 60 <0.771:0.1371 >20000 80 218.75 60 <0.76 1:0.2743

-   A: Surf. E-   B: DAZOMET-   Media: PDB-   Inoculum size: 1.156E+06-   C. albicans ATCC 102031 No synergy-   A: Surf. E-   B: DAZOMET-   Media: 1/10 TSB-   Inoculum size: 5.7E+07-   P. aeruginosa ATCC 15442 No synergy-   A: Surf. E-   B: DAZOMET-   Media: 1/10 TSB-   Inoculum size: 1.8E+06

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) S. aureus >20000 200 437.5 100 <0.52 1:0.2286 ATCC#6538

-   A: Surf. E-   B: DAZOMET-   Media: PDB-   Inoculum size: 1.156E+06

PPM AI MIC Values (3rd day) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) A. niger >20000 30 10000 2 <0.57 1:0.0002ATCC# >20000 30 10000 4 <0.63 1:0.0004 16404 >20000 30 10000 6 <0.701:0.0006 >20000 30 10000 8 <0.77 1:0.0008 >20000 30 10000 10 <0.831:0.0010 >20000 30 10000 20 <1.17 1:0.0020 >20000 30 5000 0.4 <0.261:0.0001 >20000 30 5000 0.5 <0.27 1:0.0001 >20000 30 5000 0.6 <0.271:0.0001 >20000 30 5000 0.8 <0.28 1:0.0002 >20000 30 5000 1 <0.281:0.0002 >20000 30 5000 2 <0.32 1:0.0004 >20000 30 5000 3 <0.351:0.0006 >20000 30 5000 4 <0.38 1:0.0008 >20000 30 5000 5 <0.421:0.0010 >20000 30 5000 6 <0.45 1:0.0012 >20000 30 5000 8 <0.521:0.0016 >20000 30 5000 10 <0.58 1:0.0020 >20000 30 5000 20 <0.921:0.0040 >20000 30 1750 0.4 <0.10 1:0.0002 >20000 30 1750 0.5 <0.101:0.0003 >20000 30 1750 0.6 <0.11 1:0.0003 >20000 30 1750 0.8 <0.111:0.0005 >20000 30 1750 1 <0.12 1:0.0006 >20000 30 1750 2 <0.151:0.0011 >20000 30 1750 3 <0.19 1:0.0017 >20000 30 1750 4 <0.221:0.0023 >20000 30 1750 5 <0.25 1:0.0029 >20000 30 1750 6 <0.291:0.0034 >20000 30 1750 8 <0.35 1:0.0046 >20000 30 1750 10 <0.421:0.0057 >20000 30 1750 20 <0.75 1:0.0114 >20000 30 875 3 <0.141:0.0034 >20000 30 875 4 <0.18 1:0.0046 >20000 30 875 5 <0.211:0.0057 >20000 30 875 6 <0.24 1:0.0069 >20000 30 875 8 <0.311:0.0091 >20000 30 875 10 <0.38 1:0.0114 >20000 30 875 20 <0.711:0.0229 >20000 30 437.5 0.2 <0.03 1:0.0005 >20000 30 437.5 0.3 <0.031:0.0007 >20000 30 437.5 0.4 <0.04 1:0.0009 >20000 30 437.5 0.5 <0.041:0.0011 >20000 30 437.5 0.6 <0.04 1:0.0014 >20000 30 437.5 0.8 <0.051:0.0018 >20000 30 437.5 1 <0.06 1:0.0023 >20000 30 437.5 2 <0.091:0.0046 >20000 30 437.5 3 <0.12 1:0.0069 >20000 30 437.5 4 <0.161:0.0091 >20000 30 437.5 5 <0.19 1:0.0114 >20000 30 437.5 6 <0.221:0.0137 >20000 30 437.5 8 <0.29 1:0.0183 >20000 30 437.5 10 <0.361:0.0229 >20000 30 437.5 20 <0.69 1:0.0457 >20000 30 218.75 5 <0.181:0.0229 >20000 30 218.75 6 <0.21 1:0.0274 >20000 30 218.75 8 <0.281:0.0366 >20000 30 218.75 10 <<0.34 1:0.0457 >20000 30 218.75 20 <0.681:0.0914

-   A: Surf. D-   B: DAZOMET-   Media: PDB-   Inoculum size: 1.16E+08-   P. aeruginosa ATCC 15442 No synergy-   A: Surf. D-   B: DAZOMET-   Media: PDB-   Inoculum size: 5.72E+05

PPM AI MIC Values (48 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) C. albicans >20000 50 10000 10 <0.70 1:0.0010 ATCC#10203 >20000 50 10000 20 <0.90 1:0.0020 >20000 50 10000 30 <1.101:0.0030 >20000 50 10000 40 <1.30 1:0.0040 >20000 50 5000 30 <0.851:0.0060 >20000 50 5000 40 <1.05 1:0.0080 >20000 50 2500 5 <0.231:0.0020 >20000 50 2500 10 <0.33 1:0.0040 >20000 50 2500 20 <0.531:0.0080 >20000 50 2500 30 <0.73 1:0.0120 >20000 50 2500 40 <0.931:0.0160 >20000 50 1750 6 <0.21 1:0.0034 >20000 50 1750 10 <0.291:0.0057 >20000 50 1750 20 <0.49 1:0.0114 >20000 50 1750 30 <0.691:0.0171 >20000 50 1750 40 <0.89 1:0.0229 >20000 50 875 30 <0.641:0.0343 >20000 50 875 40 <0.84 1:0.0457 >20000 50 437.5 40 <0.821:0.0914 >20000 50 218.75 40 <0.81 1:0.1829

-   A: Surf. D-   B: DAZOMET-   Media: PDB-   Inoculum size: 1.56E+06

PPM AI MIC Values (3rd day) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) A. niger >20000 40 10000 0.2 <0.51 1:0.0002ATCC# >20000 40 10000 0.3 <0.51 1:0.0003 16404 >20000 40 10000 0.4 <0.511:0.0004 >20000 40 10000 0.5 <0.51 1:0.0005 >20000 40 10000 0.6 <0.521:0.0006 >20000 40 10000 0.8 <0.52 1:0.0008 >20000 40 10000 1 <0.531:0.0010 >20000 40 10000 2 <0.55 1:0.0020 >20000 40 10000 3 <0.581:0.0030 >20000 40 10000 4 <0.60 1:0.0020 >20000 40 10000 5 <0.631:0.0030 >20000 40 10000 6 <0.65 1:0.0000 >20000 40 10000 8 <0.701:0.0008 >20000 40 10000 10 <0.75 1:0.0001 >20000 40 10000 20 <1.001:0.0001 >20000 40 5000 0.2 <0.26 1:0.0037 >20000 40 5000 0.3 <0.261:0.0046 >20000 40 5000 0.4 <0.26 1:0.0000 >20000 40 5000 0.5 <0.261:0.0001 >20000 40 5000 0.6 <0.27 1:0.0001 >20000 40 5000 0.8 <0.271:0.0002 >20000 40 5000 1 <0.28 1:0.0002 >20000 40 5000 2 <0.301:0.0004 >20000 40 5000 3 <0.33 1:0.0006 >20000 40 5000 4 <0.351:0.0008 >20000 40 5000 5 <0.38 1:0.0003 >20000 40 5000 6 <0.401:0.0010 >20000 40 5000 8 <0.45 1:0.0020 >20000 40 5000 10 <0.501:0.0030 >20000 40 5000 20 <0.75 1:0.0060 >20000 40 5000 30 <1.001:0.0020 >20000 40 2500 0.2 <0.13 1:0.0001 >20000 40 2500 0.3 <0.131:0.0001 >20000 40 2500 0.4 <0.14 1:0.0002 >20000 40 2500 0.5 <0.141:0.0002 >20000 40 2500 0.6 <0.14 1:0.0002 >20000 40 2500 0.8 <0.151:0.0003 >20000 40 2500 1 <0.15 1:0.0004 >20000 40 2500 2 <0.181:0.0008 >20000 40 2500 3 <0.20 1:0.0012 >20000 40 2500 4 <0.231:0.0016 >20000 40 2500 5 <0.25 1:0.0020 >20000 40 2500 6 <0.281:0.0024 >20000 40 2500 8 <0.33 1:0.0032 >20000 40 2500 10 <0.381:0.0040 >20000 40 2500 20 <0.63 1:0.0080 >20000 40 2500 30 <0.881:0.0120 >20000 40 1750 0.2 <0.09 1:0.0001 >20000 40 1750 0.3 <0.101:0.0002 >20000 40 1750 0.4 <0.10 1:0.0002 >20000 40 1750 0.5 <0.101:0.0003 >20000 40 1750 0.6 <0.10 1:0.0003 >20000 40 1750 0.8 <0.111:0.0005 >20000 40 1750 1 <0.11 1:0.0006 >20000 40 1750 2 <0.141:0.0011 >20000 40 1750 3 <0.16 1:0.0017 >20000 40 1750 4 <0.191:0.0023 >20000 40 1750 5 <0.21 1:0.0029 >20000 40 1750 6 <0.241:0.0034 >20000 40 1750 8 <0.29 1:0.0046 >20000 40 1750 10 <0.341:0.0057 >20000 40 1750 20 <0.59 1:0.0114 >20000 40 875 0.2 <0.051:0.0002 >20000 40 875 0.3 <0.05 1:0.0003 >20000 40 875 0.2 <0.051:0.0002 >20000 40 875 0.4 <0.05 1:0.0005 >20000 40 875 0.5 <0.061:0.0006 >20000 40 875 0.6 <0.06 1:0.0007 >20000 40 875 0.8 <0.061:0.0009 >20000 40 875 1 <0.07 1:0.0011 >20000 40 875 2 <0.091:0.0023 >20000 40 875 0.2 <0.05 1:0.0002 >20000 40 875 0.3 <0.051:0.0003 >20000 40 875 0.4 <0.05 1:0.0005 >20000 40 875 0.5 <0.061:0.0006 >20000 40 875 0.6 <0.06 1:0.0007 >20000 40 875 0.8 <0.061:0.0009 >20000 40 875 1 <0.07 1:0.0011 >20000 40 875 2 <0.091:0.0023 >20000 40 875 3 <0.12 1:0.0034 >20000 40 875 4 <0.141:0.0046 >20000 40 875 5 <0.17 1:0.0057 >20000 40 875 6 <0.191:0.0069 >20000 40 875 8 <0.24 1:0.0091 >20000 40 875 10 <0.291:0.0114 >20000 40 875 20 <0.54 1:0.0229 >20000 40 437.5 0.4 <0.031:0.0009 >20000 40 437.5 0.5 <0.03 1:0.0011 >20000 40 437.5 0.6 <0.041:0.0014 >20000 40 437.5 0.8 <0.04 1:0.0018 >20000 40 437.5 0.4 <0.031:0.0009 >20000 40 437.5 1 <0.05 1:0.0023 >20000 40 437.5 2 <0.071:0.0046 >20000 40 437.5 3 <0.10 1:0.0069 >20000 40 437.5 4 <0.121:0.0091 >20000 40 437.5 5 <0.15 1:0.0114 >20000 40 437.5 6 <0.171:0.0137 >20000 40 437.5 8 <0.22 1:0.0183 >20000 40 437.5 10 <0.271:0.0229 >20000 40 437.5 20 <0.52 1:0.0457 >20000 40 218.75 0.6 <0.031:0.0027 >20000 40 218.75 0.8 <0.03 1:0.0037 >20000 40 218.75 1 <0.041:0.0046 >20000 40 218.75 2 <0.06 1:0.0091 >20000 40 218.75 3 <0.091:0.0137 >20000 40 218.75 4 <0.11 1:0.0183 >20000 40 218.75 5 <0.141:0.0229 >20000 40 218.75 6 <0.16 1:0.0274 >20000 40 218.75 8 <0.211:0.0366 >20000 40 218.75 10 <0.26 1:0.0457 >20000 40 218.75 20 <0.511:0.0914 >20000 40 20000 0.2 <1.01 1:0.0000

-   A: Surf. D-   B: DAZOMET-   Media: 1/10 TSB-   Inoculum size: 5.72E+05

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) S. aureus >20000 200 10000 20 0.60 1:0.0020ATCC# >20000 200 10000 30 0.65 1:0.0030 6538 >20000 200 10000 40 0.701:0.0040 >20000 200 10000 50 0.75 1:0.0050 >20000 200 10000 60 0.801:0.0060 >20000 200 10000 80 0.90 1:0.0080 >20000 200 10000 100 1.001:0.0100 >20000 200 10000 20 0.60 1:0.0020 >20000 200 5000 4 0.271:0.0008 >20000 200 5000 5 0.28 1:0.0010 >20000 200 5000 6 0.281:0.0012 >20000 200 5000 8 0.29 1:0.0016 >20000 200 5000 10 0.301:0.0020 >20000 200 5000 20 0.35 1:0.0040 >20000 200 5000 30 0.401:0.0060 >20000 200 5000 8 0.29 1:0.0016 >20000 200 5000 10 0.301:0.0020 >20000 200 5000 20 0.35 1:0.0040 >20000 200 5000 30 0.401:0.0060 >20000 200 5000 40 0.45 1:0.0080 >20000 200 5000 50 0.501:0.0100 >20000 200 5000 60 0.55 1:0.0120 >20000 200 5000 80 0.651:0.0160 >20000 200 5000 100 0.75 1:0.0200 >20000 200 2500 20 0.231:0.0080 >20000 200 2500 30 0.28 1:0.0120 >20000 200 2500 40 0.331:0.0160 >20000 200 2500 50 0.38 1:0.0200 >20000 200 2500 60 0.431:0.0240 >20000 200 2500 80 0.53 1:0.0320 >20000 200 2500 100 0.631:0.0400 >20000 200 1750 0.2 0.09 1:0.0001 >20000 200 1750 0.3 0.091:0.0002 >20000 200 1750 0.4 0.09 1:0.0002 >20000 200 1750 0.5 0.091:0.0003 >20000 200 1750 0.6 0.09 1:0.0003 >20000 200 1750 0.8 0.091:0.0005 >20000 200 1750 1 0.09 1:0.0006 >20000 200 1750 2 0.101:0.0011 >20000 200 1750 3 0.10 1:0.0017 >20000 200 1750 4 0.111:0.0023 >20000 200 1750 5 0.11 1:0.0029 >20000 200 1750 6 0.121:0.0034 >20000 200 1750 8 0.13 1:0.0046 >20000 200 1750 10 0.141:0.0057 >20000 200 1750 20 0.19 1:0.0114 >20000 200 1750 40 0.291:0.0229 >20000 200 1750 60 0.39 1:0.0343 >20000 200 1750 80 0.491:0.0457 >20000 200 1750 100 0.59 1:0.0571 >20000 200 875 0.2 0.041:0.0002 >20000 200 875 0.3 0.05 1:0.0003 >20000 200 875 0.4 0.051:0.0005 >20000 200 875 0.5 0.05 1:0.0006 >20000 200 875 0.6 0.051:0.0007 >20000 200 875 0.8 0.05 1:0.0009 >20000 200 875 1 0.051:0.0011 >20000 200 875 2 0.05 1:0.0023 >20000 200 875 3 0.061:0.0034 >20000 200 875 4 0.06 1:0.0046 >20000 200 875 5 0.071:0.0057 >20000 200 875 6 0.07 1:0.0069 >20000 200 875 8 0.081:0.0091 >20000 200 875 10 0.09 1:0.0114 >20000 200 875 20 0.141:0.0229 >20000 200 875 30 0.19 1:0.0343 >20000 200 875 40 0.241:0.0457 >20000 200 875 50 0.29 1:0.0571 >20000 200 875 60 0.341:0.0686 >20000 200 875 80 0.44 1:0.0914 >20000 200 875 100 0.541:0.1143 >20000 200 437.5 30 0.17 1:0.0686 >20000 200 437.5 40 0.221:0.0914 >20000 200 437.5 50 0.27 1:0.1143 >20000 200 437.5 60 0.321:0.1371 >20000 200 437.5 80 0.42 1:0.1829 >20000 200 437.5 100 0.521:0.2286 >20000 200 218.75 50 0.26 1:0.2286 >20000 200 218.75 60 0.311:0.2743 >20000 200 218.75 80 0.41 1:0.3657 >20000 200 218.75 100 0.511:0.4571

-   A: Surf. A-   B: BIOBAN P-1487-   Media: PDB-   Inoculum size: 1.156E+06 CFU/ml

PPM AI MIC Values (3rd day) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) A. niger >20000 80 20000 5 <1.06 1:0.0003 ATCC#16404 >20000 80 10000 6 <0.58 1:0.0006 >20000 80 10000 10 <0.631:0.0010 >20000 80 10000 20 <0.75 1:0.0020 >20000 80 10000 40 <1.001:0.0040 >20000 80 5000 30 <0.63 1:0.0060 >20000 80 5000 40 <0.751:0.0080 >20000 80 5000 60 <1.00 1:0.0120 >20000 80 2500 30 <0.501:0.0120 >20000 80 2500 40 <0.63 1:0.0160 >20000 80 2500 60 <0.881:0.0240 >20000 80 1750 30 <0.46 1:0.0171 >20000 80 1750 40 <0.591:0.0229 >20000 80 1750 60 <0.84 1:0.0343 >20000 80 875 30 <0.421:0.0343 >20000 80 875 60 <0.79 1:0.0686 >20000 80 437.5 50 <0.651:0.1143 >20000 80 437.5 60 <0.77 1:0.1371 >20000 80 218.75 50 <0.641:0.2286 >20000 80 218.75 60 <0.76 1:0.2743

-   A: Surf. A-   B: BIOBAN P-1487-   Media: PDB-   Inoculum size: 1.156E+06 CFU/ml

PPM AI MIC Values (48 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) C. albicans >20000 30 20000 10 <1.33 1:0.0005 ATCC#10231 >20000 30 10000 10 <0.83 1:0.0010 >20000 30 2500 8 <0.391:0.0032 >20000 30 2500 10 <0.46 1:0.0040 >20000 30 2500 20 <0.791:0.0080 >20000 30 1750 10 <0.42 1:0.0057 >20000 30 1750 20 <0.751:0.0114 >20000 30 437.5 20 <0.69 1:0.0457 >20000 30 218.75 20 <0.681:0.0914

-   A: Surf. A-   B: BIOBAN P-1487-   Media: 1/10 TSB-   Inoculum size: 8.134E+07 CFU/ml

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) Ps. aeruginosa >20000 80 20000 60 <1.75 1:0.0030ATCC# 15442 >20000 80 5000 60 <1.00 1:0.0120 >20000 80 2500 40 <0.631:0.0160 >20000 80 2500 60 <0.88 1:0.0240 >20000 80 1750 50 <0.711:0.0286 >20000 80 1750 60 <0.84 1:0.0343 >20000 80 875 50 <0.671:0.0571 >20000 80 875 60 <0.79 1:0.0686 >20000 80 437.5 40 <0.521:0.0914 >20000 80 437.5 60 <0.77 1:0.1371 >20000 80 218.75 60 <0.761:0.2743

-   No Synergy: S. aureus-   A: Surf. E-   B: BIOBAN P-1487-   Media: PDB-   Inoculum size: 1.156E+06 CFU/ml

PPM AI MIC Values (3rd day) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) A. niger >20000 60 20000 30 <1.50 1:0.0015 ATCC#16404 >20000 60 10000 30 <1.00 1:0.0030 >20000 60 5000 30 <0.751:0.0060 >20000 60 5000 40 <0.92 1:0.0080 >20000 60 5000 50 <1.081:0.0100 >20000 60 2500 30 <0.63 1:0.0120 >20000 60 2500 40 <0.791:0.0160 >20000 60 2500 50 <0.96 1:0.0200 >20000 60 1750 30 <0.591:0.0171 >20000 60 1750 40 <0.75 1:0.0229 >20000 60 1750 50 <0.921:0.0286 >20000 60 875 30 <0.54 1:0.0343 >20000 60 875 40 <0.711:0.0457 >20000 60 875 50 <0.88 1:0.0571 >20000 60 437.5 30 <0.521:0.0686 >20000 60 437.5 40 <0.69 1:0.0914 >20000 60 437.5 50 <0.861:0.1143 >20000 60 218.75 30 <0.51 1:0.1371 >20000 60 218.75 40 <0.681:0.1829 >20000 60 218.75 50 <0.84 1:0.2286

-   A: Surf. E-   B: BIOBAN P1487-   Media: PDB-   Inoculum size: 1.808E+06

PPM AI MIC Values (48 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) C. albicans >20000 30 20000 0.04 <1.00 1:0.0000 ATCC#10203 >20000 30 10000 0.04 <0.50 1:0.0000 >20000 30 10000 0.4 <0.511:0.0000 >20000 30 10000 4 <0.63 1:0.0004 >20000 30 10000 10 <0.831:0.0010 >20000 30 10000 20 <1.17 1:0.0020 >20000 30 5000 4 <0.381:0.0008 >20000 30 5000 5 <0.42 1:0.0010 >20000 30 5000 10 <0.581:0.0020 >20000 30 5000 20 <0.92 1:0.0040 >20000 30 2500 4 <0.261:0.0016 >20000 30 2500 10 <0.46 1:0.0040 >20000 30 2500 20 <0.791:0.0080 >20000 30 1750 5 <0.25 1:0.0029 >20000 30 1750 10 <0.421:0.0057 >20000 30 1750 20 <0.75 1:0.0114 >20000 30 875 8 <0.311:0.0091 >20000 30 875 10 <0.38 1:0.0114 >20000 30 875 20 <0.711:0.0229 >20000 30 437.5 8 <0.29 1:0.0183 >20000 30 437.5 20 <0.691:0.0457 >20000 30 218.75 10 <0.34 1:0.0457 >20000 30 218.75 20 <0.681:0.0914

-   A: Surf. E-   B: BIOBAN P-1487-   Media: 1/10 TSB-   Inoculum size: 5.727E+07 CFU/ml

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) Ps. aeruginosa >20000 60 20000 50 <1.83 1:0.0025ATCC#15442 >20000 60 10000 50 <1.33 1:0.0050 >20000 60 5000 50 <1.081:0.0100 >20000 60 2500 50 <0.96 1:0.0200 >20000 60 1750 50 <0.921:0.0286 >20000 60 875 50 <0.88 1:0.0571 >20000 60 437.5 50 <0.861:0.1143 >20000 60 218.75 50 <0.84 1:0.2286

-   A: Surf. E-   B: BIOBAN P-1487-   Media: 1/10 TSB-   Inoculum size: 1.808E+06 CFU/ml

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) S. aureus >20000 30 2500 20 <0.79 1:0.0080 ATCC#6538 >20000 30 437.5 20 <0.69 1:0.0457

-   A: Surf. D-   B: BIOBAN P-1487-   Media: PDB-   Inoculum size: 1.156E+06

PPM AI MIC Values (3rd day) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) A. niger >20000 6 20000 0.6 <1.10 1:0.0000 ATCC#16404 >20000 6 10000 3 <1.00 1:0.0003 >20000 6 5000 5 <1.081:0.0010 >20000 6 2500 3 <0.63 1:0.0012 >20000 6 1750 2 <0.421:0.0011 >20000 6 875 5 <0.88 1:0.0057 >20000 6 437.5 3 <0.521:0.0069 >20000 6 218.75 4 <0.68 1:0.0183

-   A: Surf. D-   B: BIOBAN P-1487-   Media: PDB-   Inoculum size: 5.726E+05 CFU/ml

PPM AI MIC Values (48 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) C. albicans >20000 30 20000 3 <1.10 1:0.0002 ATCC#10231 >20000 30 10000 2 <0.57 1:0.0002 >20000 30 10000 3 <0.601:0.0003 >20000 30 10000 10 <0.83 1:0.0010 >20000 30 10000 20 <1.171:0.0020 >20000 30 5000 5 <0.42 1:0.0010 >20000 30 5000 10 <0.581:0.0020 >20000 30 5000 20 <0.92 1:0.0040 >20000 30 2500 0.5 <0.141:0.0002 >20000 30 2500 10 <0.46 1:0.0040 >20000 30 2500 20 <0.791:0.0080 >20000 30 1750 4 <0.22 1:0.0023 >20000 30 1750 10 <0.421:0.0057 >20000 30 1750 20 <0.75 1:0.0114 >20000 30 875 20 <0.711:0.0229 >20000 30 437.5 5 <0.19 1:0.0114 >20000 30 437.5 10 <0.361:0.0229 >20000 30 437.5 20 <0.69 1:0.0457 >20000 30 218.75 8 <0.281:0.0366 >20000 30 218.75 10 <0.34 1:0.0457 >20000 30 218.75 20 <0.681:0.0914

-   A: Surf. D-   B: BIOBAN P-1487-   Media: 1/10 TSB-   Inoculum size: 1.808E+05

PPM AI MIC Values (24 hrs) Test Alone Combination Organism CA CB Ca CbS.I. Ratio (Ca:Cb) S. aureus >20000 30 20000 20 <1.67 1:0.0010 ATCC#6538 >20000 30 10000 20 <1.17 1:0.0020 >20000 30 5000 20 <0.921:0.0040 >20000 30 2500 20 <0.79 1:0.0080 >20000 30 1750 20 <0.751:0.0114 >20000 30 218.75 20 <0.68 1:0.0914

-   No synergy: P. aeruginosa    The following biocides had no synergy against any organism tested    when paired with the following surfactants:-   Surf. A-   Sodium Benzoate, TRIS NITRO-   Surf. E-   DMDMH-   Surf. D-   CS-1246, OPP, DMDMH    In the following combinations, the ratio of surfactant to biocide    where synergy was observed were not commercially relevant, i.e., a    ratio of 1:0.2 or greater (less biocide relative to surfactant). At    these ratios, the biocide levels in a formulated product would be    too low to be practical:-   Surf. A-   DIDAC, IPBC-   Surf. E-   CMIT/MIT, IPBC, OIT, TTPC, WSCP-   Surf. D-   CMIT/MIT, OIT, DIDAC-   (MBIT, IPBC, WSCP were synergistic only at 1:0.05 or worse except    for one data point)

The invention claimed is:
 1. A synergistic microbicidal compositioncomprising: (a) a nonionic surfactant with structure:R²O(CH₂CH(CH₃)O)₃(CH₂CH₂O)₅H where R² is a mixture of C₈-C₁₄ linearalkyl groups; and (b) 2-bromo-2-nitropropane-1,3-diol; wherein a weightratio of said nonionic surfactant to 2-bromo-2-nitropropane-1,3-diol isfrom 1:0.08 to 1:0.12 or 1:0.2 to 1:1.8286.
 2. The synergisticmicrobicidal composition of claim 1 in which said mixture of C₈-C₁₄linear alkyl groups comprises from 60 to 75 wt % C₈-C₁₀ linear alkylgroups and 25 to 40 wt % C₁₂-C₁₄ linear alkyl groups.